The overall goal of the proposed project is to establish a transformation system in vitro for mammary epithelial cells that could be utilized for studies of chemical carcinogenesis at a cellular or molecular level. A new epithelial cell line, COMMA-ID, established by the principal investigator from normal mouse mammary tissue will be used in the proposed project to accomplish this goal. This cell line has advantages over previously characterized mammary cell lines in that the cells express diagnostic characteristics of normal mammary epithelial cells, (i.e. biosynthesis of casein polypeptides in vitro and ductal outgrowth in vivo that is morphologically indistinguishable from the normal mammary gland). These characteristics will permit use of this cell line for experiments relating to the following specific aims: 1. To correlate phenotypic properties in vitro of carcinogen-treated COMMA-ID cells with growth properties of the cells in vivo. 2. To establish and characterize preneoplastic and neoplastic cell lines from lesions derived from carcinogen-treated COMMA-ID cells transplanted in vivo. 3. To examine the conversion of preneoplatic cells to the tumorigenic phenotype by treatment of the cells in vitro with phorbol, a known promoting agent of mammary gland cancer in vivo. Correlations of the acquisition of transformation markers in vitro with growth potential in vivo will be made. 4. To investigate the relationship between expression of a differentiated phenotype (casein synthesis) and development of tumorigenicity. The proposed studies should provide insight into the important phenotypic alterations of mammary epithelial cells in vitro that are associated with preneoplastic and neoplastic growth in vivo.